Compositions and methods to detect glua1 in brain and to identify the presence of glua1-mediated ptsd

ABSTRACT

The present invention provides compositions and methods for detecting GluA1, as a subunit protein and/or as a GluA1-containing, GluA2-lacking AMPAR complex. The invention further provides composition and methods for detecting and/or diagnosing PTSD. The invention further relates to compositions that can be detected using radiological imaging techniques, and processes for performing such imaging techniques using the compositions, to identify elevated GluA1 expression or activity in a subject, which is a biological marker of PTSD.

RELATED APPLICATIONS

This application is a national stage entry of PCT/US17/19638, filed onFeb. 27, 2017, which claims the benefit of U.S. Provisional ApplicationNo. 62/301,583, filed on Feb. 29, 2016, the contents of each of whichare incorporated by reference in their entirety as if fully set forthherein.

BACKGROUND OF THE INVENTION

Post-Traumatic Stress Disorder (PTSD) is an incapacitating psychiatricdisorder that affects 7-10% of the U.S. population. It develops in 1 in5 people that experience or witness a traumatic event, such as warfare,natural disasters, and abuse. In a given year, about 7.7 million adults(aged 18-54) will develop PTSD symptoms and at a given time, 24.4million people have PTSD in the U.S. alone. According to the Departmentof Veterans Affairs, up to 10% of Gulf War veterans, 20% of OperationEnduring Freedom and Operation Iraqi Freedom veterans, and 30% ofVietnam War veterans have experienced PTSD symptoms. According to recentdata, PTSD imposes an annual economic burden exceeding $42 billion,mostly due to misdiagnosis and under-treatment (1).

PTSD symptoms include avoiding stimuli associated with the traumaticevent, constant re-experiencing of the event, and increased arousal,exhibited by exaggerated startle response. Under normal circumstances,these symptoms are adaptive for coping with the trauma. For instance,avoiding stimuli associated with the traumatic event lessens theprobability of encountering the threat or others like it. However,patients with PTSD lose normal daily functioning because these responsesbecome dysfunctional and exaggerated.

At present, there is no existing biological marker (or biomarker) forPTSD in humans and no objective detection systems or methods. Therefore,the only means for diagnosis of the disease are checklists of symptoms(e.g., using either the Structured Clinical Interview for DSM(SCID)—PTSD Module, or the Clinical Administered PTSD Scale (CAPS)/LifeEvents Checklist), modeled after symptoms listed in the Diagnostic andStatistical Manual for Mental Disorders (American PsychiatricAssociation, 2013) (2) (hereby incorporated by reference in its entiretyas if fully set forth herein). PTSD is underdiagnosed, partially due tothe fact that PTSD is difficult to detect with only checklists andself-report and partially because diagnoses are typically given longafter the trauma and after negative effects have manifested in thepatient.

By definition, conventional PTSD diagnosis requires manifestation ofsymptoms. The diagnostic criteria (DSM-V) include exposure to atraumatic stressor (criterion A), reexperiencing, avoidance/numbing, andhyperarousal symptoms (criteria B through D), duration of at least onemonth (criterion E), and clinically significant distress or impairmentin social/occupational functioning (criterion F) (2).

Thus, there is a need in the art for compositions and methods fordetecting and/or diagnosing PTSD, particularly at a physiological level.Relatedly, there is a need for compositions and methods of detecting inthe brain molecular increases of certain proteins known to correspond topresence of PTSD and/or cause PTSD. There is a need for detecting PTSDor its molecular causes prior to manifestation of symptoms. There isalso a need to treat PTSD once a detection and/or diagnosis has beenmade. Current treatments suppress select symptoms only and/or rely upontherapies to facilitate coping with PTSD, but a comprehensive treatmentfor PTSD that targets its physiological cause is lacking. The presentinvention satisfies these unmet needs.

SUMMARY OF THE INVENTION

The present invention is generally related to a new tracer, aradiolabeled ligand of GluA2-lacking, calcium permeable AMPA receptors(α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors orAMPARs) and/or GluA1-containing AMPARs, designed for use in radiologicalor nuclear imaging (e.g., diagnostic radiological imaging), which maydetect PTSD.

The present invention provides a method of detecting PTSD in a subject,comprising administering to the subject a first amount of a radiolabeledcomposition comprising at least one ligand of a GluA1-containing,GluA2-lacking AMPAR labeled with a radioactive isotope, wherein theradiolabeled composition comprises the following structure:

wherein the F is an [¹⁸F] radioisotope.The method further comprises creating at least one image of a brain ofthe subject using positron emission tomography (PET) or single-photonemission computed tomography (SPECT); and determining or quantifyingfrom the at least one image a GluA1-containing, GluA2-lacking AMPARdensity in the amygdala of the brain of the subject based at least inpart upon an amount of the radiolabeled composition detected in the atleast one image, and comparing the density so determined or quantifiedwith a predetermined baseline level, wherein a density greater than thepredetermined baseline level indicates PTSD in the subject.

Also provided is a method of detecting GluA1 levels, orGluA1-containing, GluA2-lacking AMPAR levels, in an amygdala of asubject, comprising administering to the subject a first amount of aradiolabeled composition comprising a ligand of GluA1 or a ligand of aGluA1-containing, GluA2-lacking AMPAR effective for detection of theradiolabeled composition in the amygdala of the subject usingradiological imaging; and determining or quantifying, by theradiological imaging of the radiolabeled composition in the amygdala ofthe subject, GluA1 subunit density or GluA1-containing, GluA2-lackingAMPAR density in the amygdala of the subject.

Also provided is a method of detecting PTSD in a subject, comprisingadministering to the subject a first amount of a radiolabeledcomposition comprising a ligand of GluA1 or a ligand of aGluA1-containing, GluA2-lacking AMPAR effective for detection of theradiolabeled composition in an amygdala of the subject usingradiological imaging; and determining or quantifying, by theradiological imaging of the radiolabeled composition in the amygdala ofthe subject, GluA1 subunit density or GluA1-containing, GluA2-lackingAMPAR density in the amygdala of the subject, and comparing the densityso determined or quantified with a predetermined baseline level, whereina density greater than the predetermined baseline level indicates PTSDin the subject.

Also provided is a method of detecting GluA1-mediated PTSD in a subject,comprising administering to the subject a first amount of a radiolabeledcomposition comprising a ligand of GluA1 or a ligand of aGluA1-containing, GluA2-lacking AMPAR effective for detection of theradiolabeled composition in an amygdala of a brain of the subject usingradiological imaging; and determining or quantifying, by theradiological imaging of the radiolabeled composition in the amygdala ofthe subject, GluA1 subunit density or GluA1-containing, GluA2-lackingAMPAR density in the amygdala of the subject, and comparing the densityso determined or quantified with a predetermined baseline level, whereina density greater than the predetermined baseline level indicatesGluA1-mediated PTSD in the subject.

The present invention also provides a method of detecting GluA1 levels(e.g., GluA1 protein levels), or GluA1-containing, GluA2-lacking AMPARlevels, in an amygdala of a subject comprising administering to thesubject a first amount of a radiolabeled composition comprising at leastone ligand of GluA1 labeled with a radioactive isotope or comprising atleast one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled witha radioactive isotope; creating at least one image of a brain of thesubject using radiological imaging (e.g., PET, SPECT, or anotherradiological detection and/or imaging system and/or device); anddetermining or quantifying from the at least one image a GluA1 subunitdensity or a GluA1-containing, GluA2-lacking AMPAR density in theamygdala of the brain of the subject based at least in part upon anamount of the radiolabeled composition detected in the at least oneimage, so as to detect GluA1 protein levels, or GluA1-containing,GluA2-lacking AMPAR levels, respectively, in an amygdala of a subject.

Also provided is a method of detecting PTSD in a subject comprisingadministering to the subject a first amount of a radiolabeledcomposition comprising at least one ligand of GluA1 labeled with aradioactive isotope or comprising at least one ligand of aGluA1-containing, GluA2-lacking AMPAR labeled with a radioactiveisotope; creating at least one image of a brain of the subject using PETor SPECT or another radiological detection and/or imaging device; anddetermining or quantifying from the at least one image a GluA1 subunitdensity or a GluA1-containing, GluA2-lacking AMPAR density in anamygdala of the brain of the subject based at least in part upon anamount of the radiolabeled composition detected in the at least oneimage. In embodiments, the method may further comprise comparing thedensity so determined or quantified with a predetermined baseline level,wherein a density greater than the predetermined baseline levelindicates PTSD in the subject. The method may further comprise providinga diagnosis for PTSD based at least in part upon a determination ofwhether the GluA1 subunit density or the GluA1-containing, GluA2-lackingAMPAR density in the amygdala exceeds the predetermined baseline levelby at least a threshold amount.

The present invention provides a method of treating PTSD in a subjectcomprising receiving information indicating a detection of elevatedlevels of GluA1, either alone as a subunit protein or inGluA1-containing, GluA2-lacking calcium permeable AMPAR complexes in anamygdala of the subject, wherein the detection has been obtained byadministering to the subject an amount of a radiolabeled compositioncomprising at least one radiolabeled ligand of GluA1 or comprising atleast one radiolabeled ligand of a GluA1-containing, GluA2-lackingAMPAR, the amount of the radiolabeled composition effective to detectthe radiolabeled composition in the amygdala of the subject usingradiological imaging (e.g., PET or SPECT); determining or quantifyingGluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in theamygdala of the subject; and determining that the GluA1 levels or theGluA1-containing, GluA2-lacking AMPAR levels exceed a predeterminedbaseline level. The method of treating further comprises administeringto the subject an amount of a treatment composition effective to treatPTSD.

In embodiments, the administered treatment composition is effective toreduce GluA1 expression levels or GluA1-containing, GluA2-lacking AMPARexpression levels in the amygdala of a subject. In embodiments, theadministered treatment composition is effective to blockGluA1-containing, GluA2-lacking AMPARs in the amygdala of a subject. Inembodiments, the administered treatment composition is effective toinhibit receptor function of GluA1-containing, GluA2-lacking AMPARs inthe amygdala of the subject.

The present invention also provides a composition comprising at leastone radiolabeled detector of a GluA1-containing, GluA2-lacking calciumpermeable (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid)receptor.

Also provided is a composition comprising at least one radiolabeleddetector of GluA1 protein.

Also provided is a compound having the following structure:

wherein the F is an [¹⁸F] radioisotope.

Also provided is a compound having the following structure:

wherein the C is a [¹¹C] radioisotope.

Also provided is a compound having the following structure:

wherein the C is a [¹¹C] radioisotope.

The present invention provides a method of producing a radiolabeledcompound comprising performing a radiofluorination reaction on a firstcompound having the following structure:

so as to produce a second compound having the following structure:

wherein the F is an [¹⁸F] radioisotope.

A method of synthesizing a radiolabeled compound is provided, whichcomprises obtaining (e.g., producing and/or trapping) an amount of[¹⁸F]; eluting the [¹⁸F] with a phase transfer catalyst KF/K2.2.2 so asto produce a solution of [¹⁸F]KF/K2.2.2 complex; adding a first compoundhaving the following structure:

to the solution of [¹⁸F]KF/K2.2.2 complex so as to perform aradiofluorination reaction to create a second compound having thefollowing structure:

wherein the F is an [¹⁸F] radioisotope. In embodiments, the method mayfurther comprise purifying the reaction contents from to produce apurified compound radiolabeled with [¹⁸F]. In embodiments, purifying thereaction contents can comprise performing radio-HPLC on the product ofthe radiofluorination reaction.

BRIEF DESCRIPTION OF THE DRAWINGS

The following detailed description of preferred embodiments of theinvention will be better understood when read in conjunction with theappended drawings. For the purpose of illustrating the invention, thereare shown in the drawings embodiments which are presently preferred. Itshould be understood, however, that the invention is not limited to theprecise arrangements and instrumentalities of the embodiments shown inthe drawings.

FIG. 1A illustrates an experimental design for Stress-Enhanced FearLearning (SEFL), a rodent model of PTSD.

FIGS. 1B-D depict representative Western blot images and graphs ofrelative optical density ratios (±standard error of the mean (SEM)) ofGluA1, GluA2, and GluN1, respectively, and a glyceraldehyde 3-phosphatedehydrogenase (GAPDH) control. These figures show the results of exampleexperiments demonstrating that there are observed long-term increases inGluA1 protein in the basolateral amygdala (BLA) after a traumatic event.Glutamate receptor protein changes were measured three weeks after theinitial trauma via Western blotting. While the GluA2 subunit of theAMPAR and the GluN1 subunit of the N-methyl-D-aspartate receptor (NMDAR)remained unchanged after trauma, GluA1 increased substantially.Metyrapone, a cortisol/corticosterone-synthesis blocker given before thetrauma effectively preventing SEFL also prevented GluA1 increases andconferred levels to that of unstressed controls.

FIG. 2A illustrates an abbreviated SEFL experimental design.

FIG. 2B illustrates cannulae placement into the basolateral amygdala(BLA) of rodents.

FIGS. 2C-E depict the results of example experiments demonstrating thatintra-BLA infusions of GluA1 antisense oligonucleotide (ASO) after the15-shock stressor reverse the SEFL. Delivering GluA1 ASO post-traumasignificantly decreased GluA1 protein levels in stressed rats. Missenseoligonucleotide (MSO) control infusions still conferred high level ofGluA1 protein, as determined by Western blotting (FIGS. 2C-D) GluA1 ASOpost-trauma also prevented the sensitized fear (i.e., freezing)typically observed in novel Context B after just a mild shock (labeledin the graph as the conditioning context) after receiving a trauma incontext A (FIG. 2E).

FIG. 3A illustrates a SEFL experimental design.

FIG. 3B illustrates cannulae placement verification in the BLA.

FIG. 3C depicts the results of example experiments demonstrating thatintra-BLA infusions of IEM-1460, a selective GluA2-lacking AMPARantagonist, after the 15-shock stressor attenuate SEFL. BlockingGluA2-lacking AMPARs in the amygdala post-trauma block the sensitizedfear (i.e., freezing) typically observed in novel Context B after just amild shock (labeled in the graph as the conditioning context) afterreceiving a trauma in Context A.

FIG. 4 illustrates a topographical model of channel binding sites inAMPARs. Hydrophobic and nucleophilic regions in the receptor areseparated by approximately 10 A. Therefore, only compounds possessing a‘head and tail’ structure such as IEM-1460 can block the channel.

FIG. 5 is a flowchart illustrating an exemplary process for synthesizinga radiolabeled composition in exemplary embodiments of the presentinvention.

DETAILED DESCRIPTION

The present invention relates generally to compositions and methods fordetecting levels (e.g., densities) of GluA1 (also referred to as GluR1),either alone as a subunit protein or within GluA1-containing,GluA2-lacking AMPAR complexes in an amygdala of a subject (e.g., a humansubject). In embodiments, the compositions and methods of the presentinvention can enable detection and/or diagnosis of PTSD in a subject.The compositions and methods may comprise use of imaging techniques(e.g., diagnostic and/or radiological imaging techniques) involvingimaging agents designed to target GluA1 and/or GluA1-containing and/orGluA2-lacking AMPARs in the brain.

AMPARs are composed of four types of protein subunits, which aredesignated as GluA1 GluA2 (also referred to as GluR2), GluA3 (alsoreferred to as GluR3), and GluA4 (also referred to as GluR4) (3). TheHUGO gene nomenclature committee refers to the encoding gene for GluA1as HGNC:4571, and for GluA2 as HGNC:4572. EauA1 is at times referred toby the gene name, GRIM, which produces GluA1 protein when translation isactivated (e.g., by a learning event). The GRIA2 gene produces GluA2protein when translation is activated. Non-limiting exemplary GRIMsequences include NCBI Reference Sequences: NM000827.3; NM001258023.1and NM001258022.1. Non-limiting exemplary GRIA2 sequences include NCBIReference Sequences: NM000826.3, NM001083620.1, and NM001083619.1.

The AMPAR subunits combine to form tetramers, protein polymers composedof four monomer units. The presence of a GluA2 subunit will almostalways render the receptor channel impermeable to calcium, meaning thatthe neurons on which these AMPARs are located are more difficult todepolarize, transduce signals, and/or communicate with neighboringneurons. Without cell activity, GluA2/GluA3-containing AMPARs are morecommonly located at the synapse because of their stability to betethered to the cell membrane. On the other hand, GluA1-containingAMPARs are important for cell plasticity and are brought to cellsynapses in the presence of cell activity to strengthen cellconnections. GluA1-containing AMPARs that are lacking GluA2 are commonlyfound in areas important for learning and memory and particularly fearlearning and memory, such as the hippocampus and the amygdala.Enrichment of synaptic GluA2-lacking AMPARs (presumably GluA1 homomers),as well as synaptic insertion of GluA1 in these regions underlieslong-term potentiation (LTP), a process by which recent patterns ofactivity cause persistent strengthening of synapses, which is crucialfor long-term memory formation. Blocking the formation of GluA1 proteinwill effectively block the formation of functional GluA1-containingAMPARs after a learning event and/or potentially block LTP/long-termmemory formation (4).

A composition is provided comprising at least one radiolabeled detectorof GluA1, either alone as a subunit protein or as part of aGluA1-containing, GluA2-lacking calcium permeable(α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor complex,which may be a human AMPAR. Accordingly the composition may comprise atleast one radiolabeled detector of a GluA1-containing, GluA2-lackingcalcium permeable AMPAR. The radiolabeled detector may comprise a ligandof a GluA1-containing, GluA2-lacking calcium permeable AMPAR. Inembodiments, a composition may comprise at least one radiolabeleddetector of GluA1 protein, e.g., as a subunit protein. In embodiments,the radiolabeled detector may be a detector of GluA1 expression, surfaceexpression, AMPAR activity, or a combination thereof. In embodiments,the radiolabeled detector may be an inhibitor of AMPAR function. Inembodiments, such an inhibitor may be an inhibitor of calcium permeableAMPAR function.

In embodiments, the radiolabeled detector may be selected from the groupconsisting of a nucleic acid, an antisense nucleic acid (e.g., GluA1ASO), a ribozyme, a peptide, a small molecule (e.g., a molecule of anorganic compound having a low molecular weight (e.g., less than 900daltons) that may help regulate a biological process, with a size on theorder of 1 nm), an antagonist, an aptamer, and a peptidomimetic. Theradiolabeled detector may comprise a radioisotope, such as [¹⁸F] or[¹¹C].

In embodiments, the radiolabeled detector may compriseN′-[(4-fluoro-1-adamantyl)methyl]pentane-1,5-diamine. In embodiments,the radiolabeled detector may comprise the following structure:

wherein the F is an [¹⁸F] isotope.

In embodiments, the radiolabeled detector may comprise the followingstructure:

wherein the C is a [¹¹C] isotope.

In embodiments, the radiolabeled detector may comprise the followingstructure:

wherein the C is a [¹¹C] isotope.

In embodiments, the composition may comprise a solution of theradiolabeled detector, such as a saline solution and/or an ethanolsolution. In embodiments, a solution comprising the radiolabeleddetector may be diluted, e.g., with water or saline. The composition maybe put into a solution, e.g., for administration.

In embodiments, the composition can serve as a radiolabeled tracer in animaging detection process, e.g., radiological or nuclear imaging, suchas PET, SPECT, or other radiological sensing and/or imaging, such asusing a portable sensing device. In embodiments, the imaging detectionprocess may be used to quantify and/or determine expression (e.g.,levels, amounts, and/or densities) of GluA1 protein or ofGluA1-containing, GluA2-lacking AMPARs in an amygdala of a subject. Inembodiments, amygdala refers to both a left amygdala and a rightamygdala together. In embodiments, only a portion of the amygdala isstudied, imaged, and/or examined, such as the left portion or the rightportion, which may include one or more nuclei on either side of theamygdala, such as the BLA. Accordingly, a BLA may be examined, which maybe a left BLA, a right BLA, or both a left and a right BLA. Inembodiments, the subject is a human subject. In embodiments, the imagingdetection process may be used to to determine whether the GluA1 proteinitself or the GluA1-containing, GluA2-lacking AMPARs exceed apredetermined baseline level. In embodiments, the imaging detectionprocess may be used to determine a degree to which and/or an amount bywhich the GluA1 protein or the GluA1-containing, GluA2-lacking AMPARsexceed a predetermined baseline level. In embodiments, the imagingdetection process may be usable to detect PTSD in a subject, e.g., basedon detected levels (e.g., densities) of GluA1, either alone as a subunitprotein or in GluA1-containing, GluA2-lacking calcium permeable AMPARcomplexes in an amygdala of a subject, and/or further based upon acomparison to one or more baseline levels (e.g., baseline densities) orthreshold levels.

In embodiments, a baseline or predetermined baseline level (e.g., ofGluA1 levels or of GluA1-containing, GluA2-lacking AMPAR levels (e.g.,density)) may be determined by performing the imaging detection processon a plurality of subjects known not to be suffering from PTSD;determining respective amygdalar GluA1 levels or GluA1-containing,GluA2-lacking AMPAR levels, in each of the plurality of subjects; andcomputing as the predetermined baseline level a normalized average ofthe respective amygdalar GluA1 levels or GluA1-containing, GluA2-lackingAMPAR levels in each of the plurality of subjects.

In embodiments, the predetermined baseline level (e.g., of GluA1 levels(e.g., density) or of GluA1-containing, GluA2-lacking AMPAR levels(e.g., density)) may be determined by performing the imaging detectionprocess on a first plurality of subjects known not to be suffering fromPTSD; determining first respective amygdalar GluA1 density orGluA1-containing, GluA2-lacking AMPAR density in each of the firstplurality of subjects; computing a first normalized average of the firstrespective amygdalar GluA1 density or GluA1-containing, GluA2-lackingAMPAR density; performing the imaging detection process on a secondplurality of subjects known to be suffering from PTSD; determiningsecond respective amygdalar GluA1 density or GluA1-containing,GluA2-lacking AMPAR density in each of the second plurality of subjects;computing a second normalized average of the second respective amygdalarGluA1 density or GluA1-containing, GluA2-lacking AMPAR density; anddetermining as the predetermined baseline level an amygdalar GluA1density or GluA1-containing, GluA2-lacking AMPAR density based at leastin part upon the first normalized average and the second normalizedaverage. Determining the baseline may comprise performing one or morestudy replications and/or comparing across many subjects, e.g., usingstatistical analyses.

A compound is provided having the following structure:

wherein the F is an F-18 radioisotope.

A compound is provided having the following structure:

wherein the C is a [¹¹C] radioisotope.

A compound is provided having the following structure:

wherein the C is a [¹¹C] radioisotope.

The present invention provides a method of detecting PTSD in a subject,comprising administering to the subject a first amount of a radiolabeledcomposition comprising at least one ligand of a GluA1-containing,GluA2-lacking AMPAR labeled with a radioactive isotope, wherein theradiolabeled composition comprises the following structure:

wherein the F is an [¹⁸F] radioisotope.The method further comprises creating at least one image of a brain ofthe subject using radiological imaging (e.g., PET or SPECT); anddetermining or quantifying from the at least one image aGluA1-containing, GluA2-lacking AMPAR density in the amygdala of thebrain of the subject based at least in part upon an amount of theradiolabeled composition detected in the at least one image, andcomparing the density so determined or quantified with a predeterminedbaseline level, wherein a density greater than the predeterminedbaseline level indicates PTSD in the subject.

A method of detecting GluA1, either alone as a subunit protein or inGluA1-containing, GluA2-lacking calcium permeable AMPAR complexes in theamygdala of a subject is provided. In embodiments, the method ofdetecting GluA1 (e.g., GluA1 levels and/or GluA1 subunit density) orGluA1-containing, GluA2-lacking AMPARs (e.g., GluA1-containing,GluA2-lacking AMPAR levels and/or density) may comprise administering tothe subject a first amount of a radiolabeled composition (e.g., a traceror radiotracer) comprising a ligand of GluA1 or a ligand of aGluA1-containing, GluA2-lacking AMPAR effective for detection of theradiolabeled composition in the amygdala of the subject usingradiological imaging (e.g., PET or SPECT); and determining orquantifying, by the radiological imaging of the radiolabeled compositionin the amygdala of the subject, GluA1 subunit density orGluA1-containing, GluA2-lacking AMPAR density in the amygdala of thesubject.

A method of detecting PTSD (e.g., GluA1-mediated PTSD) in a subject isprovided. The method may comprise administering to the subject a firstamount of a radiolabeled composition comprising a ligand of GluA1 or aligand of a GluA1-containing, GluA2-lacking AMPAR effective fordetection of the radiolabeled composition in an amygdala of a brain ofthe subject using radiological imaging; and determining or quantifying,by the radiological imaging of the radiolabeled composition in theamygdala of the subject, GluA1 subunit density or GluA1-containing,GluA2-lacking AMPAR density in the amygdala of the subject, andcomparing the density so determined or quantified with a predeterminedbaseline level, wherein a density greater than the predeterminedbaseline level indicates PTSD (e.g., GluA1-mediated PTSD) in thesubject. In embodiments, such an evaluation using the detection methodsof the present invention may be performed after a subject experiences atrauma and/or before manifestation of symptoms (e.g., observablesymptoms). In embodiments, the subject is a subject who experienced atrauma and/or a subject being evaluated for presence of and/or severityof PTSD. In embodiments, evaluation for PTSD may be performed before,during, and/or after treatment, and/or multiple times during the courseof a treatment regimen.

In embodiments, the ligand is a ligand of GluA1. In such cases, theprocess may determine GluA1 levels, such as GluA1 subunit density. Inembodiments, the ligand is a ligand of a GluA1-containing, GluA2-lackingAMPAR. In such cases, the process may determine GluA1-containing,GluA2-lacking AMPAR levels and/or density. In embodiments, theradiological imaging is PET or SPECT imaging. In embodiments, theradiological imaging comprises using another radiation sensing,detection, and/or imaging device. Such a device may be a portabledevice, such as a handheld electronic device. Accordingly, theradiological imaging may comprise using a portable electronic device todetect radiation levels associated with the radiolabeled composition(e.g., associated with an amount of the radiolabeled composition thathas undergone uptake and accumulation in the amygdala). In embodiments,such a device may comprise a cellular phone and/or an integrated camera.The radiation sensing device may have installed thereon onnon-transitory computer-readable memory or otherwise be operable withparticularly programmed software, which can evaluate detected radiationlevels, e.g., to provide an indication or notification of such levelsand/or to provide an indication or notification that such levels exceeda preprogrammed baseline level. Such software may comprise aninstallable application, such as a downloadable or an uploadableapplication.

In embodiments, determining or quantifying GluA1 subunit density and/orlevels of GluA1-containing, GluA2-lacking AMPAR density and/or levels inthe amygdala of the subject can comprise determining or quantifying anamount of the radiolabeled composition in the amygdala afteradministration. In embodiments, determining or quantifying GluA1 subunitdensity or GluA1-containing, GluA2-lacking AMPAR density in the amygdalaof the subject can comprise detecting, determining, quantifying,visualizing, and/or estimating radioactive emissions of the radiolabeledcomposition in the amygdala after administration. Detectable and/ormeasurable radioactive emissions may provide a proxy for detectingand/or measuring GluA1 itself or GluA1-containing, GluA2-lacking AMPARs,via receptor binding of the ligands. Accordingly, radioactive emissionsmay estimate GluA1 levels and/or GluA1-containing/GluA2-lacking AMPARdensity, according to the respective ligand used. Radioactive emissionsof the administered radiolabeled composition may be detected, quantifiedvisualized, and/or estimated following an uptake period of time after orduring which the composition can accumulate and/or bind to its targetGluA1 protein or GluA1-containing, GluA2-lacking AMPARs. Suchradioactive emissions from the radiolabeled composition or a portionthereof (e.g., the portion that bound to targets in an area of interestand/or area to be imaged) may be detected by a special-purpose camera orimaging device that can produce pictures, provide molecular information,and/or provide indications of detected radiation levels, e.g.,indicating the locations of such detected radiation.

In embodiments, the method may further comprise comparing theradiolabeled composition in the amygdala after administration to acontrol amount. In embodiments, the method may further comprisecomparing the radioactive emissions of the radiolabeled composition inthe amygdala after administration to a control (e.g., a control image, acontrol amount or control level or control density, or a control densityrepresentation, to name a few). In embodiments, the method may comprisecomparing receptor density associated with the radioactive emissions toa control receptor density. In embodiments, a control densityrepresentation may comprise a visual representation of a control amountof GluA1 or of GluA1-containing, GluA2-lacking AMPARs in an amygdala,which may be a particular subject's amygdala (e.g., with a visualrepresentation of the control amount overlaid thereon) or an artisticrendering of an amygdala with the control amount. Such a visualrepresentation may be generated (e.g., by particularly programmedsoftware) and/or provided to technicians, doctors, and/or otherpersonnel, and/or used for diagnostic purposes.

The control amount and/or the control receptor density may be apredetermined baseline level, which baseline may be determined asdescribed herein. In embodiments, the baseline level may be associatedwith a particular radiolabeled composition. In embodiments, the controlamount may be an amount or estimation of radioactive emissions from anamygdala. In embodiments, the control amount may be an amount, level,and/or density of GluA1 protein or of GluA1-containing, GluA2-lackingAMPARs. In embodiments, the method may further comprise comparing theemissions of the radiolabeled composition in the amygdala afteradministration to a reference brain region or other organ with a knownor approximately known GluA1 amount or GluA1-containing, GluA2-lackingAMPAR amount. In embodiments, the method may comprise using an uptakevalue, such as a standardized uptake value and/or a fractional uptakevalue, to determine levels of GluA1, either alone as a subunit proteinor in GluA1-containing, GluA2-lacking calcium permeable AMPAR complexes.

A method of detecting GluA1, either itself (e.g., detecting the GluA1protein subunit itself, whether isolated or not isolated from othercompositions) in an amygdala of a subject or as part of aGluA1-containing, GluA2-lacking AMPAR complex in the amygdala of thesubject is provided. In embodiments, the method is a method of detectingGluA1 protein levels, or of GluA1-containing, GluA2-lacking AMPARlevels, in an amygdala of a subject. In embodiments, the method maydetermine or detect elevated GluA1 levels and/or that GluA1 levels arenot elevated (e.g., at or around baseline or below baseline). The methodcomprises administering to the subject a first amount of a radiolabeledcomposition comprising at least one ligand of GluA1 labeled with aradioactive isotope or comprising at least one ligand of aGluA1-containing, GluA2-lacking AMPAR labeled with a radioactiveisotope; creating at least one image of a brain of the subject or aregion thereof (e.g., the amygdala, such as the left amygdala and/or theright amygdala) using radiological imaging (e.g., PET or SPECT); anddetermining or quantifying from the at least one image a GluA1 level(e.g., GluA1 subunit density) or GluA1-containing, GluA2-lacking AMPARlevel (e.g., density level) in the amygdala of the brain of the subjectbased at least in part upon an amount of the radiolabeled composition(e.g., an amount of selective uptake and accumulation of theradiolabeled composition in the brain) detected and/or detectable in theat least one image and/or an amount of radioactive emissions detected.

Accordingly, the method may comprise administering to the subject afirst amount of a radiolabeled composition comprising at least oneligand of GluA1 labeled with a radioactive isotope or comprising atleast one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled witha radioactive isotope; creating at least one image of a brain of thesubject using PET or SPECT; and determining or quantifying from the atleast one image a GluA1 subunit density or a GluA1-containing,GluA2-lacking AMPAR density in the amygdala of the brain of the subjectbased at least in part upon an amount of the radiolabeled compositiondetected in the at least one image, e.g., so as to detect GluA1 proteinlevels, or GluA1-containing, GluA2-lacking AMPAR levels, respectively,in an amygdala of a subject.

The present invention provides a method of detecting PTSD (e.g.,GluA1-mediated PTSD) in a subject, comprising administering to thesubject a first amount of a radiolabeled composition comprising at leastone ligand of GluA1 labeled with a radioactive isotope or comprising atleast one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled witha radioactive isotope; creating at least one image of a brain of thesubject using PET or SPECT or another radiological detection and/orimaging device; and determining or quantifying from the at least oneimage a GluA1 subunit density or a GluA1-containing, GluA2-lacking AMPARdensity in an amygdala of the brain of the subject based at least inpart upon an amount of the radiolabeled composition detected in the atleast one image. In embodiments, the method may further comprisecomparing the density so determined or quantified with a predeterminedbaseline level, wherein a density greater than the predeterminedbaseline level indicates PTSD (e.g., GluA1-mediated PTSD) in thesubject.

In embodiments, the radiolabeled composition can serve as an imagingagent composition, e.g., to enable and/or to facilitate radiologicalimaging, such as via PET or SPECT. In embodiments, the amount of theradiolabeled composition detected and/or detectable in the at least oneimage comprises an amount of radioactive emissions detected, e.g.,detected in the amygdala. Accordingly, the methods of detection providedherein may comprise determining or quantifying from the at least oneimage a GluA1 level or GluA1-containing, GluA2-lacking AMPAR level inthe amygdala of the brain of the subject based at least in part upon adetected amount of radioactive emissions, e.g., from the amygdala. Theradioactive emissions correspond to a portion of the first amount of theradiolabeled composition that has undergone selective uptake andaccumulation, e.g., by binding to GluA1 itself or to GluA1-containing,GluA2-lacking AMPARs, respectively.

In embodiments, the methods of detection may be performed and/or thefirst amount of the radiolabeled composition may be administered to thesubject at least one hour after the subject experiences a trauma (e.g.,a physical trauma and/or a mental trauma) and/or many years after thetrauma, e.g., for as long as PTSD symptoms persist. Each patient may begiven more than one scan (e.g., using the detection methods of thepresent invention) during his or her lifetime. In embodiments, the atleast one image of the brain may be created between 15 minutes and 3hours following administration of the radiolabeled composition.

In embodiments, the methods of detection may further comprisedetermining whether GluA1, either alone as a subunit protein or as partof a GluA1-containing, GluA2-lacking AMPAR complex in the amygdalaexceeds a predetermined baseline level, which baseline may be determinedas described herein. In embodiments, the methods may further compriseproviding a diagnosis for PTSD based at least in part upon thedetermination of whether GluA1 levels (e.g., GluA1 subunit density) orGluA1-containing, GluA2-lacking AMPAR levels (e.g., density) in theamygdala exceeds the predetermined baseline level by at least athreshold amount. In embodiments, the threshold amount is zero, and thebaseline is a GluA1-containing, GluA2-lacking AMPAR level over which thesubject has elevated levels of GluA1-containing, GluA2-lacking AMPARscorresponding to PTSD levels. In embodiments, the threshold amount maybe a percentage amount greater than a baseline level of non-PTSDsubjects, which percentage amount may fall in the range of 5-10%, 5-15%,or 5-25%, to name a few. In embodiments, the threshold amount may be anumeric amount of GluA1-containing, GluA2-lacking AMPARs greater than abaseline level of non-PTSD subjects. The threshold for indicating,determining, and/or diagnosing PTSD may be determined by comparison(e.g., statistical analysis) of measured levels of GluA1-containing,GluA2-lacking AMPAR in respective amygdalae of control subjects and ofsubjects suffering from PTSD.

A method of treating PTSD in a subject is provided. The method comprisesreceiving information indicating a detection of elevated levels ofGluA1, either alone as a subunit protein or in GluA1-containing,GluA2-lacking calcium permeable AMPAR complexes in an amygdala of thesubject, wherein the detection has been obtained by administering to thesubject an amount of a radiolabeled composition comprising at least oneradiolabeled ligand of GluA1 or comprising at least one radiolabeledligand of a GluA1-containing, GluA2-lacking AMPAR, the amount of theradiolabeled composition effective to detect the radiolabeledcomposition in the amygdala of the subject using radiological imaging(e.g., PET or SPECT); determining or quantifying GluA1 levels orGluA1-containing, GluA2-lacking AMPAR levels in the amygdala of thesubject; and determining that the GluA1 levels or the GluA1-containing,GluA2-lacking AMPAR levels exceed a predetermined baseline level, whichmay be determined as described herein. The method further comprisesadministering to the subject an amount of a treatment compositioneffective to treat PTSD.

In embodiments, the treatment composition comprises a GluA1-containing,GluA2-lacking AMPAR ligand. In embodiments, the treatment compositioncomprises an inhibitor of calcium permeable AMPAR function.

In embodiments, the administered treatment composition (e.g., therespective amount of the treatment composition) is effective to reduceGluA1 expression levels (e.g., thereby blocking the formation offunctional GluA1-containing AMPARs) or GluA1-containing, GluA2-lackingAMPAR expression levels in the amygdala of a subject. In embodiments,the administered treatment composition is effective to blockGluA1-containing, GluA2-lacking AMPARs in the amygdala of the subject.In embodiments, such a treatment composition may be selected from thegroup consisting of a nucleic acid, an antisense nucleic acid, aribozyme, a peptide, a small molecule, an inhibitor of GluA1 expressionor synthesis, an aptamer, and a peptidomimetic.

In embodiments, the respective amount of the treatment composition iseffective to inhibit receptor function of GluA1-containing,GluA2-lacking AMPARs in the amygdala of the subject. In embodiments,such a treatment composition may be selected from the group consistingof a peptide, a small molecule, an antagonist, an inhibitor, and apeptidomimetic. In embodiments, such a treatment composition mayalleviate one or more symptoms of PTSD.

In embodiments, the radiolabeled composition is a radiolabeled detectoror otherwise may have any of the properties and/or structures asdescribed herein with respect to radiolabeled detectors. Theradiolabeled composition may comprise a saline and/or ethanol solution.

In embodiments, the radiolabeled composition may be administered to thesubject via injection into the bloodstream (e.g., intravenous injectionand/or intravenous drip), via injection into tissue and/or an organ, viaenema, orally (e.g., in pill, tablet, or liquid form), via inhalation,and/or via a nasal spray, in a manner effective to enter the amygdala ofthe subject.

In embodiments, the amount of the radiolabeled composition administeredto the subject (e.g., the first amount) comprises a mass dose of theradiolabeled composition in a range of 0.0001 pg to 1 ng per kg of bodyweight of the subject. In embodiments, such a mass dose may fall in therange 0.01-2 pg/kg. In embodiments, an amount of radioactivityassociated with the amount of the radiolabeled composition administeredto the subject falls in a range of 1 to 2000 MBq. In embodiments, such aradioactivity level may fall in a range of 1-150 MBq, 100-250 MBq,200-300 MBq, 150-370 MBq, 300-400 MBq, or 400-2000 MBq, to name a few.In some embodiments an effective radiation dose equivalent associatedwith the amount of the radiolabeled composition administered to thesubject falls in a range of 1 to 100 μSv, which is linearly related toradioactivity. In embodiments, the radiation dosages may fall in a rangeof 1-12 μSv, 10-25 μSv, 20-30 μSv, 30-40 μSv, to name a few.

Administration of the radiolabeled composition (and/or a solutionthereof) in accordance with the present invention may occur in one bolusadministered some time (e.g., an uptake period) before the detectionand/or imaging takes place. However, in embodiments, the administrationof the agents of the invention may be essentially continuous over apreselected period of time, may comprise in a series of spaced doses, ormay comprise a singly administered dose, depending on factors known toskilled practitioners. Both local and systemic administration arecontemplated. The amount administered may vary depending on variousfactors including, but not limited to, the composition chosen, aparticular target being evaluated (e.g., GluA1 protein orGluA1-containing, GluA2-lacking AMPARs), a particular disease beingevaluated (e.g., PTSD, anxiety, to name a few), the weight, the physicalcondition, and/or the age of the subject. Such factors can be determinedby a clinician employing animal models or other test systems which arewell known to the art. An effective amount of a composition for use inan imaging diagnostic procedure may be an amount sufficient to bedetected by the imaging procedure, e.g., PET technique via detection ofradioactive emissions, or may be an amount sufficient to bind to atarget receptor.

When the radiolabeled ligands of the invention are prepared foradministration, they may be combined with a pharmaceutically acceptablecarrier, diluent, or excipient to form a pharmaceutical formulation, orunit dosage form.

In embodiments, an uptake period during which the radiolabeledcomposition travels through a body of a subject may be in the range of 5to 100 minutes, 45 to 60 minutes, or 45 to 90 minutes, to name a few.Accordingly, an uptake period may be 5 minutes, 15 minutes, 30 minutes,45 minutes, 50 minutes, 60 minutes, 80 minutes, 85 minutes, or 90minutes, to name a few. An image acquisition time or a scan duration fora radiological imaging procedure (e.g., a PET scan or a SPECT scan) maybe in the ranges of 5 to 45 minutes, 15 to 60 minutes, 30 to 60 minutes,or 30 to 90 minutes, to name a few, such as scan durations of 5 minutes,10 minutes, 15 minutes, 20 minutes, or 30 minutes, to name a few. Inembodiments, shorter scan durations may be performed, such as using aportable radiological sensing and/or imaging device.

In certain aspects the present invention relates to detecting anincrease in the expression, surface expression, and/or activity ofGluA1, an AMPAR subunit, by using specially designed imaging agentcompositions. In certain embodiments, the method relates to detectingthe activity of GluA2-lacking, calcium-permeable AMPARs, which areprimarily composed of GluA1 subunits.

In one aspect of the present invention, a composition is created byradiolabeling a ligand for use with radiological imaging (e.g., PET orSPECT) or other diagnostic imaging systems and processes, the specificligand being designed to bind with GluA1-containing and/or GluA2-lackingAMPARs, preferentially in an amygdala of a subject's brain.

In embodiments, the present invention comprises a method for diagnosingPTSD in a subject using a radiolabeled composition that is a ligand fora particular biomarker as an imaging agent (e.g., diagnostic imagingagent) and/or tracer for determining presence of and/or quantifyinglevels of the biomarker. The invention utilizes radiological (e.g., PETor SPECT) imaging, a technique that allows for the identification ofmolecular and cellular levels, such as brain receptor levels (e.g.,density), in living humans. The technique may enable determination ofchanges in such levels. The imaging process uses small amounts ofradiolabeled compounds (e.g., radiolabeled with positron-emittingisotopes, such as [¹⁸F]fluorine, [¹²⁴I]iodine, or [¹¹C]carbon) havinghigh specificity for the target molecule, chemical, protein, or othertarget of interest; radiolabeled compounds having high affinity (e.g.,in pM and/or nM range) are needed for such measurements. The inventioncomprises at least one ligand of GluA1-containing and/or calciumpermeable AMPARs labeled with a radioactive isotope. PET scanning orother radiological detection and/or imaging of a subject using thiscomposition can reveal sites of high receptor concentration (e.g., AMPARdensity). In embodiments, such sites can include the amygdala in thebrain.

In embodiments, the diagnostic imaging agent is a ligand of AMPARfunction. In embodiments, the ligand is an inhibitor of GluA2-lacking,calcium permeable and/or GluA1-containing AMPAR function. Inembodiments, the composition comprises IEM-1460 (e.g., radiolabeled witha radioisotope such as [¹¹C]). In embodiments, the composition comprisesIEM-1925 dihydrobromide. In embodiments, the composition comprisesphilanthotoxin 74. In embodiments, the composition comprises Naspmtrihydrobromide. In embodiments, the ligand is an agonist ofGluA2-lacking, calcium permeable and/or GluA1-containing AMPAR function.In embodiments, the composition comprises Cl-HIBO.

In embodiments, the present invention provides a composition fordetecting PTSD in a subject. In embodiments, the composition comprises aradiolabeled ligand of GluA2-lacking, calcium permeable AMPARs and/orGluA1-containing AMPARs. In embodiments, the composition is a diagnosticimaging agent. In embodiments, the composition comprises a radiolabeledligand of AMPAR function. In embodiments, the ligand is a radiolabeledinhibitor of calcium permeable and/or GluA1-containing AMPAR function.In embodiments, the ligand is a radiolabeled agonist of calciumpermeable and/or GluA1-containing AMPAR function. In embodiments, theradiolabeled inhibitor is an inhibitor of calcium-permeable AMPAR, suchas IEM-1460. In embodiments, the radiolabeled ligand is an agonist ofcalcium permeable AMPARs, such as Cl-HIBO.

In embodiments, the present invention provides a method for diagnosingPTSD. In embodiments, the method comprises administering a ligand of aGluA1-containing and/or GluA2-lacking AMPAR labeled with a radioactiveisotope to a subject in a diagnostic imaging setting and detectinglevels of GluA1-containing and/or GluA2-lacking AMPARs by proxy ofradioactive emissions from the ligand labeled with the radioactiveisotope. In certain embodiments, the composition comprises a ligand ofGluA1 expression, surface expression, activity, or a combination thereofthat has been labeled with a radiolabeled isotope to detect expressionlevels in the brain. These compounds may be radiolabeled withpositron-emitting isotopes, including but not limited to [¹⁸F]fluorine,[¹²⁴I]iodine, [¹¹C]carbon, [¹⁵O]oxygen, [¹³N]nitrogen, or [⁷⁶Br]bromide,using organic chemistry and/or radiochemistry methods.

In embodiments, a method for detecting PTSD can comprise administeringto a human subject (e.g., orally or via intravenous injection) animaging agent composition comprising at least one ligand of GluA1labeled with a radioactive isotope. Such a composition is not found innature and is markedly different from naturally occurring molecules. Theimaging agent composition may be designed to bind with GluA1-containingand/or GluA2-lacking AMPARs. The method can further comprise creating atleast one image of the brain of the human subject using PET or SPECT, orany other radiological imaging method; determining from the at least oneimage a GluA1 receptor density (which may be a density range) in theamygdala of the brain of the human subject based at least in part uponan amount of the imaging agent composition detectable in the at leastone image; determining whether the GluA1-containing and/or GluA2-lackingAMPAR density in the amygdala exceeds a predefined baseline level ofGluA1; and providing a diagnosis for PTSD based at least in part uponthe determination of whether the GluA1-containing and/or GluA2-lackingAMPAR density in the amygdala exceeds the predefined baseline level byat least a threshold amount.

In certain embodiments, the method comprises detecting GluA1-containingand/or GluA2-lacking, calcium permeable AMPARs specifically in the leftand/or right BLA of the subject. In certain embodiments, the methodcomprises administering a composition that is directed or targeted tothe BLA.

In certain embodiments, the method comprises administering thecomposition to a subject who may have PTSD or is at risk for developingPTSD. For example, in certain embodiments, the method comprisesadministering the radiolabeled ligand and performing PET or SPECT scanimaging on a subject who has experienced a traumatic event. Exemplarytraumatic events include but are not limited to, violence, assault,military experiences, accidents or near-accidents, natural disasters,and the like. In embodiments, the detection method is a diagnosticmethod. In embodiments, the diagnostic is performed within a definedperiod after the traumatic event. For example, in certain embodiments,the radiolabeled ligand is administered and PET scan is performed withinminutes, hours, days (e.g., 1-3 days), weeks, months, or years after thetraumatic event.

In embodiments, the present invention provides a composition fordiagnosing PTSD in a subject. For example, in certain embodiments, thecomposition is used to detect PTSD in a subject that has experienced atrauma.

In embodiments, the composition comprises a ligand of and/or inhibitorof GluA1-containing and/or GluA2-lacking, calcium permeable AMPARs. Forexample, in embodiments, the composition inhibits GluA1 expression,GluA1 surface expression, GluA1 activity, or a combination thereof. Inembodiments, the ligand and/or inhibitor is radiolabeled with aradioactive isotope.

In embodiments, the composition inhibits the activity of GluA1 includingGluA1-containing AMPARs. In embodiments, the inhibitor inhibits theactivity of GluA2-lacking, calcium permeable AMPARs. In certainembodiments, the inhibitor inhibits ionic influx through the AMPAR;inhibits agonist binding of the AMPAR; inhibits pore opening of theAMPAR; enhances pore blocking of the AMPAR; inhibits calcium influx ofthe AMPAR; or a combination thereof.

An inhibitor of GluA1 is any compound, molecule, or agent that reduces,inhibits, or prevents the function of GluA1. For example, an inhibitorof GluA1 is any compound, molecule, or agent that reduces GluA1expression, surface expression, activity, or a combination thereof. Incertain embodiments, the inhibitor inhibits the transcription of DNA,inhibits the translation of RNA, and/or inhibits the protein itself. Inembodiments, an inhibitor of GluA1 comprises a peptide, an antibody, asmall molecule, a ribozyme, an antagonist, an aptamer, a peptidomimetic,or any combination thereof.

In embodiments, the composition comprises an agonist of GluA1-containingand/or GluA2-lacking calcium permeable AMPARs. For example, in oneembodiment, the composition activates GluA1 expression, GluA1 surfaceexpression, GluA1 activity, or a combination thereof.

In embodiments, the composition stimulates the activity of GluA1including GluA1-containing AMPARs. In embodiments, the agonist activatesthe activity of GluA2-lacking, calcium permeable AMPARs. In certainembodiments, the agonist increases ionic influx through the AMPAR;inhibits antagonist binding of the AMPAR; increases pore opening of theAMPAR; reduces pore blocking of the AMPAR; increases calcium influx ofthe AMPAR; or a combination thereof.

An agonist of GluA1 is any compound, molecule, or agent that increasesor stimulates the function of GluA1. For example, an agonist of GluA1 isany compound, molecule, or agent that increases GluA1 expression,surface expression, activity, or a combination thereof. In certainembodiments, the agonist increases the transcription of DNA, increasesthe translation of RNA, or activates the protein itself. In embodiments,an agonist of GluA1 comprises a peptide, an antibody, a small molecule,a ribozyme, an antagonist, an aptamer, a peptidomimetic, or anycombination thereof.

The present invention relates generally to compositions and methods fordiagnosing PTSD in a subject using diagnostic imaging techniques. Incertain aspects the present invention relates to detecting an increasein the expression, surface expression, and/or activity of GluA1, anAMPAR subunit. In certain embodiments, the method relates to detectingthe activity of GluA2-lacking calcium permeable AMPARs, which areprimarily composed of GluA1 subunits.

In one aspect, the present invention provides a method for diagnosingPTSD in a subject, comprising administering at least one radiolabeledinhibitor of GluA1-containing or calcium permeable AMPARs and detectingGluA1-containing or calcium permeable AMPARs, e.g., via radioactiveemissions.

In embodiments, the present invention provides a method for diagnosingPTSD. In embodiments, the method comprises administering a ligand of aGluA1-containing or GluA2-lacking, calcium permeable AMPAR labeled witha radioactive isotope to a subject in a diagnostic imaging setting andmeasuring radioactive emissions from a portion of a brain (e.g., theamygdala) of the subject. For example, in certain embodiments, acomposition that is administered to a subject comprises a ligand ofGluA1 expression, surface expression, activity, or a combination thereofthat has been labeled with a radiolabeled isotope to detect expressionlevels in the brain. In embodiments, the composition comprises a ligandof a GluA1-containing or calcium permeable AMPAR labeled with aradioactive isotope. In certain embodiments, the method comprisesdetecting GluA1-containing or GluA2-lacking, calcium permeable AMPARsspecifically in the BLA of the subject. In certain embodiments, themethod comprises administering a composition that is directed ortargeted to the BLA.

In embodiments, the present invention provides a method for diagnosingPTSD by performing a PET scan using a radiolabeled GluA1-containingand/or GluA2-lacking, calcium permeable AMPAR ligand in a subject whohas experienced a trauma. In embodiments, the method comprisesdiagnosing PTSD by performing a PET scan using a radiolabeledGluA1-containing and/or GluA2-lacking, calcium permeable AMPAR ligand toa subject having a genetic predisposition to developing PTSD. Inembodiments, the method comprises diagnosing PTSD by performing a PETscan using a radiolabeled GluA1-containing and/or GluA2-lacking, calciumpermeable AMPAR ligand to a subject who has experienced a traumaticevent. In certain embodiments, the subject has been the victim of, or awitness to, a traumatic event, including, but not limited to, violence,assault, sexual assault, accident, vehicle accident, near accident,natural disaster, military violence, or the like.

In certain embodiments, the method comprises diagnosing PTSD byperforming a PET scan using a radiolabeled GluA1-containing and/orGluA2-lacking, calcium permeable AMPAR ligand in a defined time periodfollowing the traumatic event. For example, in certain embodiments, themethod comprises diagnosing PTSD by performing a PET scan using aradiolabeled GluA1-containing and/or GluA2-lacking, calcium permeableAMPAR ligand within 1 day, 2 days, 3 days, 5 days, 7 days, 10 days, 2weeks, 4 weeks, 1 month, 2 months, 3 months, 6 months, 9 months, 1 year,2 years, 5 years, or 10 years following the traumatic event, to name afew. The detection methods of the invention may be used to diagnose PTSDin any subject. In embodiments, the subject is a mammal, including, butnot limited to, a human, primate, cow, horse, sheep, goat, dog, cat,rodent, and the like.

Labeling a GluA1-containing and/or GluA2-lacking, calcium permeableAMPAR ligand with a radioactive isotope can be done using any methodknown to the skilled artisan. Performing a PET scan using a radiolabeledGluA1-containing and/or GluA2-lacking, calcium permeable AMPAR ligandcan be done using any method known to the skilled artisan. A GluA1ligand may therefore be a compound that inhibits or activates GluA1function, activity, or stability. A GluA1 ligand may be any type ofcompound, including but not limited to, peptide, a small molecule, orcombinations thereof. GluA1-containing and/or GluA2-lacking, calciumpermeable AMPA receptor specific binding may be accomplished eitherdirectly or indirectly. Methods of decreasing or increasing expressionof GluA1 include, but are not limited to, methods that use a ribozyme, apeptide, a small molecule, and combinations thereof.

Administration of a radiolabeled GluA1-containing or GluA2-lacking,permeable AMPAR ligand and detecting radioactive emissions therefrom canbe used along with a treatment or treatment regimen to determinesubjects requiring treatment, track treatment progress, and/or determinewhen treatment is no longer needed.

In embodiments, a radiolabeled GluA1-containing and/or GluA2-lacking,calcium permeable AMPAR ligand is administered to a subject, andradioactive emissions are detected and/or quantified. The ligand mayalso be a hybrid or fusion composition to facilitate, for instance,delivery to target cells or efficacy. In embodiments, a hybridcomposition may comprise a tissue-specific targeting sequence. Forexample, in embodiments, the ligand is targeted to the BLA of thesubject.

EXAMPLES Introduction

Rodent experiments have suggested a link between upregulation of GluA1in the amygdala of a subject and PTSD. Using a rodent model of PTSDcalled SEFL, it has been shown that after a traumatic event, there areenduring increases in GluA1 protein in the BLA (5, 6) (herebyincorporated by reference in their entireties as if fully set forthherein). However, other glutamate receptor subunits, such as the GluA1subunit of the AMPAR and GluN1 subunit of the NMDAR, do not show anylong-term changes (FIGS. 1A-D).

FIG. 1A illustrates an experimental design using SEFL (7, 8). Proteinlevels were measured using Western blot, an analytical technique todetect specific proteins in tissue samples.

FIG. 1B depicts representative Western blot images of GluA1 and acontrol, GAPDH, from the BLA of stressed and unstressed rats, measuredthree weeks after the trauma. The graph shows mean GluA1: GAPDH opticaldensity ratios (±SEM). GluA1 protein levels in the BLA weresignificantly higher in stressed rats than in unstressed rats andmetyrapone-treated rats; (**p<0.01, two-way analysis of variance(ANOVA), followed by a priori planned comparisons) (5).

FIG. 1C depicts representative Western blot images of GluA2 and GAPDHfrom the BLA of stressed and unstressed rats, measured three weeks afterthe trauma. The graph shows mean GluA2: GAPDH optical density ratios(±SEM). Neither stress nor metyrapone had an effect on GluA2 levels (5).

FIG. 1D depicts representative Western blot images of GluN1 and GAPDHfrom the BLA of stressed and unstressed rats, measured three weeks afterthe trauma. The graph shows mean GluN1: GAPDH optical density ratios(±SEM). Neither stress nor metyrapone had an effect on GluN1 levels (5).

Because GluA1 increases are observed weeks after the trauma, post-traumaGluA1 ASO infusions directly into the BLA prevented sensitized fearresponses usually observed in SEFL (FIGS. 2A-E) (5). FIG. 2A illustratesthe experimental design, and FIG. 2B shows the cannulae placement. FIG.2C depicts representative Western blot images of GluA1 and GAPDH fromthe BLA of stressed/MSO and stressed/ASO rats. FIG. 2D depicts meanGluA1: GAPDH optical density ratios (±SEM) from the BLA of stressed/MSOand stressed/ASO rats. ASO significantly lowers GluA1 levels in the BLAwhen delivered after the trauma compared with MSO (*p<0.05, one-wayANOVA). FIG. 2E depicts mean (±SEM) percent freezing in Context B on Day3. GluA1 ASO significantly reduces conditional freezing compared withstressed controls (i.e., those infused with artificial cerebrospinalfluid (ACSF) or MSO) when delivered into the BLA post-trauma tounstressed levels (* p<0.05, one-way ANOVA followed by plannedcomparisons).

Similar results were observed using IEM-1460, a GluA2-lacking,GluA1-containing AMPAR antagonist, suggesting that the GluA1 subunitincrease also increased viable GluA1-containing AMPARs to act upon(FIGS. 3A-C) (5). FIG. 3A depicts a SEFL experimental design, and FIG.3B shows cannulae placement verification. FIG. 3C depicts mean freezing(±SEM) in Context B on Day 3. IEM-1460 infused post-trauma significantlyreduces freezing in Context B compared with stressed rats infused withACSF and unstressed controls (* p<0.05, one-way ANOVA, followed by apriori planned comparisons) (5).

This work discussed hereinabove was conducted in rodents using Westernblotting techniques to detect glutamate receptor protein levels. Westernblotting to detect specific proteins in humans is not viable in livingsubjects; it requires using post-mortem tissue and artificial antibodiesto react with the target protein in this tissue. The present inventionprovides alternative methods to detect GluA1 protein levels utilizingnuclear imaging, which can be performed in live human patients. Insteadof designing antibodies to react with the target protein in the tissuesample, nuclear imaging requires injecting a radioisotope withspecificity to the protein that will also be able to penetrate theregion of interest in awake patients. In particular for the presentinvention, nuclear imaging can be effected by radiolabeling a ligand ofGluA1 or of GluA1-containing, GluA2-lacking AMPARs, such as those usedin the outlined studies. While IEM-1460 is a known ligand ofGluA1-containing, GluA2-lacking AMPARs, radiolabeling this compounddirectly with [¹⁸F] in a manner that preserves its ability to bind toGluA1 is not feasible.

Difficulty of Radiolabeling

The most direct extrapolation of the example studies would be toradiolabel an already existing and commercially available drug thatbinds to GluA1-containing/GluA2-lacking AMPARs, such as IEM-1460(N,N,H-Trimethyl-5-[(tricyclo[3.3.1.13,7]dec-1-ylmethyl)amino]-1-pentanaminiumbromidehydrobromide). IEM-1460 or any other GluA2-lacking AMPAR ligands havenever before been radiolabeled.

Radiolabeling such a ligand is complicated by the necessity to add theradioisotope at a particular location of the structure so as not tointerfere with and/or prevent binding. FIG. 4 illustrates exemplaryAMPAR channel binding sites (9). Ligands of AMPARs have similar ‘headand tail’ structures with two primary structural features. Thequarternary amine on the alkyl chain 402 is thought to associate withthe pore region of ionotropic channels as a cation mimic. The adamantaneportion 404 of the molecule is a hydrophobic moiety that fills space inthe pore and confers the steric hindrance that prevents cation passagethrough the pore. According to Bolshakov et al., the terminal ammoniumgroup is important for binding; and the chain length is crucial to allowbinding to calcium permeable AMPARs (9). Chain length between adamantanemoiety and terminal ammonium should be approximately 10 Å so that theadamantane can interact with the hydrophobic region at the pore of thereceptor; while the ammonium group can interact with the nucleophilicregion of the receptor.

In embodiments, [¹¹C] labeling of IEM-1460 may be possible. However, the[¹⁸F] radioisotope is preferred to [¹¹C] for radiolabeling because itshalf-life of about 110 minutes is much greater than the approximately20-minute half-life of [¹¹C]. Because of its short half-life, hasseverely limited clinical usability, requiring an on-site cyclotron toperform the labeling (10). There are some radiometals suitable forclinical use (e.g., [⁶⁸Ga]) but these are connected to the tracer vialarge/complex chelators and this would impact the pharmacologicproperties of the product rendering it unusable for labeling AMPARligands.

A strategy for [¹⁸F] labeling has not previously been discussed orproposed. IEM-1460 does not contain any halogen atoms. Directfluorination of the product would most likely target the nucleophilicalkyl amine chain. Interfering with the alkyl amine would likely affectchannel pharmacology by altering the nitrogen basicity. While IEM-1460has been fluorinated with [¹⁹F] in the past, the conditions used are notcompatible with [¹⁸F] radiolabeling (11, 12).

Given that direct fluorination of IEM-1460 is unfavorable and/orinfeasible in terms of preserving its ability and/or specificity to bindto AMPARs, incorporating [¹⁸F] into the compound or otherwise creating aradiolabeled ligand of AMPARs and in particular GluA2-lacking AMPARspresents technical challenges.

Method of Manufacture

A radiolabeling process was developed to produce a radiolabeled compoundthat is a ligand of GluA1 or of GluA1-containing, GluA2-lacking AMPARswith a radiolabel a radioisotope) in a location that does not preventbinding to the AMPARs. In the synthesis methods disclosed herein, asdepicted in FIG. 5, a modified adamantane derivative 502 is selectedensuring the amine tail of the molecule satisfied a length correspondingto AMPAR binding. The specific binding properties of the precursorcompound 502 with AMPARs were not previously known. While adamantanederivatives are known generally to bind to calcium permeable AMPARs,compound 502 had not yet been studied for those purposes and has onlyseldomly been used in scientific research. Moreover, adamantanederivatives in general are not common to radiolabel with [¹⁸F] becausethere are no halogen-group atoms to replace in the molecule. Aradiochemistry leaving group, tosylate, is used on the far end ofadamantane in compound 502, to facilitate novel [¹⁸F] radiolabeling atthis structure instead of at the tail.

As depicted in FIG. 5, the present invention provides a method ofproducing a radiolabeled compound comprising performing aradiofluorination reaction on a first compound 502 having the followingstructure:

wherein OTs is tosyl and Boc is tert-Butyloxycarbonyl, so as to producea second compound 504 having the following structure:

wherein the F is an [¹⁸F] radioisotope.

In embodiments, compound 502 is[5-[[5-(tert-butoxycarbonylamino)pentylamino]methyl]-2-adamantyl]4-methylbenzenesulfonateor (1R,3 S,5 s,7s)-5-(((5-((tert-butoxycarbonyl)amino)pentyl)amino)methyl)adamantan-2-yl4-methylbenzenesulfonate.

In embodiments, compound 504 isN′-[(4-fluoro-1-adamantyl)methyl]pentane-1,5-diamine or N1-(((1 s,3R,5S,7 s)-4-fluoroadamantan-1-yl)methyl)pentane-1,5-diaminium chloride.

In embodiments, the present invention provides a method of synthesizingand/or producing a radiolabeled compound. The method comprisessynthesizing an [¹⁸F]fluoride solution comprising [¹⁸F]fluoride via a(p,n) reaction of [¹⁸O]water by proton bombardment in a cyclotron. Inembodiments to synthesize a compound radiolabeled with a [¹¹C]radioisotope, this step may comprise synthesizing [¹¹C]carbon dioxidevia a (p,n) reaction of [¹¹B] by proton bombardment in a cyclotron. Themethod of producing the radiolabeled compound further comprisesextracting and/or trapping the [¹⁸F]fluoride by passing the[¹⁸F]fluoride solution through a preconditioned QMA anion exchangecartridge; eluting and/or heating, in a reaction vessel, the extracted[¹⁸F]fluoride with a first solvent comprising an acetonitrile(MeCN)/water solution containing a phase transfer catalyst (e.g.,potassium carbonate (K2CO3) and/or Kryptofix 2.2.2 (KF/K2.2.2); applyinga vacuum to the reaction vessel so as to remove the first solvent andleave a dried residue of a [¹⁸F]KF/K2.2.2 complex; adding, to thereaction vessel, MeCN to re-dissolve the dried residue; evaporating, viaapplication of heat and vacuum to the reaction vessel, so as to removeresidual water via azeotropic distillation leaving a second driedresidue of [¹⁸F]KF/K2.2.2 complex; mixing, in a second solvent (e.g.,approximately 1 mL of a second solvent, such as MeCN or dimethylsulfoxide (DMSO)) in the reaction vessel, the second dried residue of[¹⁸F]KF/K2.2.2 complex and a compound 502 having the structure shown inFIG. 5; heating the reaction vessel so as to perform a radiofluorinationreaction to produce a radiolabeled compound 504 having the followingstructure shown in FIG. 5, wherein the F is an [¹⁸F] radioisotope; andpurifying, via radio-High Performance Liquid Chromatography (HPLC), theradiofluorination reaction contents to produce a purified radiolabeledcompound. In embodiments, the reaction vessel may comprise one or moredifferent reaction vessels to which reaction contents are added. Heatinga reaction vessel heats such reaction contents.

HPLC is an analytical chemistry technique used to separate, identify,and/or quantify each component in a mixture, which technique is usablehere to evaluate and/or ensure purity of the radiolabeled composition.In embodiments, the method may further comprise formulating the purifiedradiolabeled compound in saline to prepare for human use. Purificationusing HPLC may be performed to prepare a solution for administration. Inembodiments, the resulting purified fraction may then be reformulatedfor administration (e.g., injection) either by evaporating off all thesolvent and replacing with saline or flowing the pure fraction through asep-pak cartridge (e.g., C18) to trap the compound. The cartridge canthen be washed with water, and the compound may be eluted off thecartridge in a low amount of ethanol. The eluted compound may be dilutedwith saline, e.g., such that ethanol is <10% v/v. The result may bepassed through a sterile filter. In embodiments, quality control testingmay be performed.

In embodiments of the present invention, a method of synthesizing aradiolabeled compound comprises obtaining (e.g., producing and/ortrapping) an amount of [¹⁸F]; eluting the [¹⁸F] with a phase transfercatalyst KF/K2.2.2 so as to produce a solution of [¹⁸F]KF/K2.2.2complex; adding a first compound 502 having the structure shown in FIG.5 to the solution of [¹⁸F]KF/K2.2.2 complex (e.g., approximately 1 mL ofthe solution) so as to perform a radiofluorination reaction to create asecond compound 504 having the structure shown in FIG. 5, wherein the Fis an [¹⁸F] radioisotope. In embodiments, performing theradiofluorination comprises adding heat to the reaction contents and/orto a reaction vessel containing such reaction contents. In embodiments,the method may further comprise purifying the reaction contents from toproduce a purified compound radiolabeled with [¹⁸F]. In embodiments,purifying the reaction contents can comprise performing radio-HPLC onthe product of the radiofluorination reaction.

In embodiments, a method of synthesizing a radiolabeled compoundcomprises isolating [¹⁸F]; combining the isolated [¹⁸F] with a phasetransfer catalyst (e.g., KF/K2.2.2) to produce an [¹⁸F] phase transferbuffer; combining that complex with compound 502 having the structureillustrated in FIG. 5; and adding heat to produce compound 504.

Definitions

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as commonly understood by one of ordinary skill inthe art to which this invention belongs. Although any methods andmaterials similar or equivalent to those described herein can be used inthe practice or testing of the present invention, the preferred methodsand materials are described.

The nomenclature used herein and the laboratory procedures used inanalytical chemistry, radiochemistry, and organic syntheses describedherein are those well known and commonly employed in the art. Standardtechniques or modifications thereof, are used for chemical syntheses andchemical analyses.

As used herein, each of the following terms has the meaning associatedwith it in this section.

The articles “a” and “an” are used herein to refer to one or to morethan one (i.e., to at least one) of the grammatical object of thearticle. By way of example, “an element” means one element or more thanone element.

“About” as used herein when referring to a measurable value such as anamount, a temporal duration, and the like, is meant to encompassvariations of ±20% or ±10%, more preferably ±5%, even more preferably±1%, and still more preferably ±0.1% from the specified value, as suchvariations are appropriate to perform the disclosed methods.

“Agonist” refers to a chemical that binds to a receptor and activatesthe receptor to produce a biological response. The agonist can beendogenous, coming from within the body, or exogenous, coming fromoutside the body, such as a drug.

“Antagonist” refers to a chemical or drug that blocks or dampensagonist-mediated responses rather than provoking a biological responseitself upon binding to a receptor. They are sometimes called blockers;examples include calcium channel blockers. In pharmacology, antagonistshave affinity but no efficacy for their receptors to which they bind,and binding will disrupt the interaction and inhibit the function of anagonist.

“Antisense” refers particularly to the nucleic acid sequence of thenon-coding strand of a double stranded DNA molecule encoding a protein,or to a sequence which is substantially homologous to the non-codingstrand. As defined herein, an antisense sequence is complementary to thesequence of a double stranded DNA molecule encoding a protein. It is notnecessary that the antisense sequence be complementary solely to thecoding portion of the coding strand of the DNA molecule. The antisensesequence may be complementary to regulatory sequences specified on thecoding strand of a DNA molecule encoding a protein, which regulatorysequences control expression of the coding sequences. “Complementary” asused herein refers to the broad concept of subunit sequencecomplementarity between two nucleic acids, e.g., two DNA molecules. Whena nucleotide position in both of the molecules is occupied bynucleotides normally capable of base pairing with each other, then thenucleic acids are considered to be complementary to each other at thisposition. Thus, two nucleic acids are substantially complementary toeach other when at least about 50%, preferably at least about 60% andmore preferably at least about 80% of corresponding positions in each ofthe molecules are occupied by nucleotides which normally base pair witheach other (e.g., A:T and G:C nucleotide pairs).

“Biomarker” refers to a measurable substance in an organism whosepresence is indicative of some phenomenon such as disease, infection, orenvironmental exposure.

As used herein, “aptamer” refers to a small molecule that can bindspecifically to another molecule. Aptamers are typically eitherpolynucleotide- or peptide-based molecules. A polynucleotide aptamer isa DNA or RNA molecule that adopts a highly specific three-dimensionalconformation designed to have appropriate binding affinities andspecificities towards specific target molecules, such as peptides,proteins, drugs, vitamins, among other organic and inorganic molecules.Such polynucleotide aptamers can be selected from a vast population ofrandom sequences through the use of systematic evolution of ligands byexponential enrichment. A peptide aptamer is typically a loop of about10 to about 20 amino acids attached to a protein scaffold that binds tospecific ligands. Peptide aptamers may be identified and isolated fromcombinatorial libraries, using methods such as the yeast two-hybridsystem.

The terms “diagnose,” “diagnosing,” and “diagnosis,” refer to detectingmeasures described herein. The methods of “diagnosis” can employ usingdetection and/or imaging techniques, such as PET scan imaging, on asubject to identify target compounds in target anatomical regions, toidentify presence of and/or quantities of target compounds in targetanatomical regions, to identify characteristics of target compounds intarget anatomical regions, and/or to identify characteristics of targetanatomical regions. This can require administration of a composition ofthe present invention, for example, a subject possibly afflicted with adisease or disorder, in order to detect, identify, determine theseverity of, or determine the course of treatment of the disorder orrecurring disorder, or in order to prolong the survival of a subjectbeyond that expected in the absence of such diagnosis.

A “disease” is a state of health of an animal wherein the animal cannotmaintain homeostasis, and wherein if the disease is not ameliorated thenthe animal's health continues to deteriorate. In contrast, a “disorder”in an animal is a state of health in which the animal is able tomaintain homeostasis, but in which the animal's state of health is lessfavorable than it would be in the absence of the disorder. Leftuntreated, a disorder does not necessarily cause a further decrease inthe animal's state of health.

A disease or disorder is “alleviated” if the severity or frequency of atleast one sign or symptom of the disease or disorder experienced by apatient is reduced.

“Encoding” refers to the inherent property of specific sequences ofnucleotides in a polynucleotide, such as a gene, a cDNA, or an mRNA, toserve as templates for synthesis of other polymers and macromolecules inbiological processes having either a defined sequence of nucleotides(i.e., rRNA, tRNA and mRNA) or a defined sequence of amino acids and thebiological properties resulting therefrom. Thus, a gene encodes aprotein if transcription and translation of mRNA corresponding to thatgene produces the protein in a cell or other biological system. Both thecoding strand, the nucleotide sequence of which is identical to the mRNAsequence and is usually provided in sequence listings, and thenon-coding strand, used as the template for transcription of a gene orcDNA, can be referred to as encoding the protein or other product ofthat gene or cDNA.

The terms “effective amount” and “pharmaceutically effective amount”refer to a nontoxic but sufficient amount of an agent to provide thedesired biological result. That result can be reduction and/oralleviation of the signs, symptoms, or causes of a disease or disorder,or any other desired alteration of a biological system. An appropriateeffective amount in any individual case may be determined by one ofordinary skill in the art using routine experimentation.

As used herein “endogenous” refers to any material from or producedinside an organism, cell, tissue or system.

As used herein, the term “exogenous” refers to any material introducedfrom or produced outside an organism, cell, tissue or system.

The term “expression” as used herein is defined as the transcriptionand/or translation of a particular nucleotide sequence driven by itspromoter.

The term “isotope” refers to any variant of a particular chemicalelement which differ in neutron number, although all isotopes of a givenelement have the same number of protons in each atom. For example,carbon-12, carbon-13 and carbon-14 are three isotopes of the elementcarbon with mass numbers 12, 13 and 14 respectively. “Radiolabeled” or“Radioactive” as applied to an object, sometimes called “radioisotope”refers to a composition that has excess nuclear energy, making itunstable. This excess energy can either create and emit, from thenucleus, new radiation (gamma radiation) or a new particle (alphaparticle or beta particle), or transfer this excess energy to one of itselectrons, causing it to be ejected (conversion electron). During thisprocess, the object is said to undergo radioactive decay. Radioisotopesare used for diagnosis, treatment, and research. Radioactive chemicaltracers emitting gamma rays or positrons can provide diagnosticinformation about internal anatomy and the functioning of specificorgans. Radioisotopes can be attached to a ligand in order to determinereceptor binding. This is used in some forms of tomography such as PETscanning.

The term “ligand” refers to any substance that forms a complex with abiomolecule, and can serve a biological purpose. In protein-ligandbinding, the ligand is usually a molecule which produces a signal bybinding to a site on a target protein (usually a receptor). The bindingtypically results in a change of conformation of the target protein. InDNA-ligand binding studies, the ligand can be a small molecule, ion orprotein, that binds to a particular part of the DNA double helix. Therelationship between ligand and binding partner is a function of charge,hydrophobicity, and molecular structure. In terms of ligand-receptorbinding, the ligand can either be an agonist or antagonist (competitiveor non-competitive) of the receptor. “Radiolabeled” or “Radioactive” asapplied to an object, sometimes called “radioligand” or “tracer” refersto a biochemical substance (in particular, a ligand that isradiolabeled) that is used for diagnosis or for research-oriented studyof the receptor systems of the body. In a neuroimaging application theradioligand can be injected into the pertinent tissue, or infused intothe bloodstream and binds to its receptor. In embodiments, theradioligand may be administered orally via swallowing, by inhalation, byinjection (intravenous), and/or by enema. When the radioactive isotopein the ligand decays, it can be measured by PET or SPECT scan imaging.It is often used to quantify the binding of a test molecule to thebinding site of radioligand.

By “nucleic acid” is meant any nucleic acid, whether composed ofdeoxyribonucleosides or ribonucleosides, and whether composed ofphosphodiester linkages or modified linkages such as phosphotriester,phosphoramidate, siloxane, carbonate, carboxymethylester, acetamidate,carbamate, thioether, bridged phosphoramidate, bridged methylenephosphonate, phosphorothioate, methylphosphonate, phosphorodithioate,bridged phosphorothioate or sulfone linkages, and combinations of suchlinkages. The term nucleic acid also specifically includes nucleic acidscomposed of bases other than the five biologically occurring bases(adenine, guanine, thymine, cytosine and uracil). The term “nucleicacid” typically refers to large polynucleotides.

The term “(p,n) reaction” refers to a type of nuclear reaction thatoccurs when a neutron enters a nucleus and a proton leaves the nucleussimultaneously.

“Polypeptide” refers to a polymer composed of amino acid residues,related naturally occurring structural variants, and syntheticnon-naturally occurring analogs thereof linked via peptide bonds.Synthetic polypeptides can be synthesized, for example, using anautomated polypeptide synthesizer.

The term “protein” typically refers to large polypeptides.

The term “peptide” typically refers to short polypeptides. In anembodiment, a peptide is 20 amino acids or less in length. In anembodiment, a peptide is 10 amino acids or less in length.

As used herein, a “peptidomimetic” is a compound containing non-peptidicstructural elements that is capable of mimicking the biological actionof a parent peptide. A peptidomimetic may or may not comprise peptidebonds.

The term “oligonucleotide” typically refers to short polynucleotides,generally no greater than about 60 nucleotides. It will be understoodthat when a nucleotide sequence is represented by a DNA sequence (i.e.,A, T, G, C), this also includes an RNA sequence (i.e., A, U, G, C) inwhich “U” replaces “T.” “Recombinant polynucleotide” refers to apolynucleotide having sequences that are not naturally joined together.An amplified or assembled recombinant polynucleotide may be included ina suitable vector, and the vector can be used to transform a suitablehost cell.

The term “positron emission tomography (PET)” refers to a functionalimaging technique that is used to observe metabolic processes in thebody. The system detects pairs of gamma rays emitted indirectly by apositron-emitting radioisotope (tracer), which is introduced into thebody on a biologically active molecule.

The term “single-photon emission computed tomography (SPECT)” is anuclear medicine tomographic imaging technique that is used to observemetabolic processes in the body. SPECT is similar to PET in its use ofradioactive tracer material and detection of gamma rays. In contrastwith PET, however, the tracers used in SPECT emit gamma radiation thatis measured directly, whereas PET tracers emit positrons that annihilatewith electrons up to a few millimeters away, causing two gamma photonsto be emitted in opposite directions.

By the term “specifically binds,” as used herein, is meant a molecule,such as a ligand, which recognizes and binds to another molecule orfeature, such as a receptor, but does not substantially recognize orbind other molecules or features in a sample.

The phrase “inhibit,” as used herein, means to reduce a molecule, areaction, an interaction, a gene, an mRNA, and/or a protein'sexpression, stability, function or activity by a measurable amount or toprevent entirely. Inhibitors are compounds that, e.g., bind to,partially or totally block stimulation, decrease, prevent, delayactivation, inactivate, desensitize, or down regulate a protein, a gene,and an mRNA stability, expression, function and activity, e.g.,antagonists. Inhibiting a receptor means reducing a parameter of thereceptor's function(s).

“Therapeutically effective amount” is an amount of a compound of theinvention, that when administered to a patient, ameliorates a symptom ofthe disease. The amount of a compound of the invention which constitutesa “therapeutically effective amount” will vary depending on thecompound, the disease state and its severity, the age of the patient tobe treated, and the like. The therapeutically effective amount can bedetermined routinely by one of ordinary skill in the art having regardto his own knowledge and to this disclosure.

The terms “subject” or “patient” for the purposes of the presentinvention includes humans and other animals, particularly mammals, andother organisms. Thus the methods are applicable to both human therapyand veterinary applications. In a preferred embodiment the patient is amammal, and in a most preferred embodiment the patient is human.

The terms “treat,” “treating,” and “treatment,” refer to therapeutic orpreventative measures described herein. The methods of “treatment”employ administration to a subject, in need of such treatment, acomposition of the present invention, for example, a subject afflicted adisease or disorder, or a subject who ultimately may acquire such adisease or disorder, in order to prevent, cure, delay, reduce theseverity of, or ameliorate one or more symptoms of the disorder orrecurring disorder, or in order to prolong the survival of a subjectbeyond that expected in the absence of such treatment.

Ranges: throughout this disclosure, various aspects of the invention canbe presented in a range format. It should be understood that thedescription in range format is merely for convenience and brevity andshould not be construed as an inflexible limitation on the scope of theinvention. Accordingly, the description of a range should be consideredto have specifically disclosed all the possible subranges as well asindividual numerical values within that range. For example, descriptionof a range such as from 1 to 6 should be considered to have specificallydisclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numberswithin that range, for example, 1, 2, 2.7, 3, 4, 5, 5.3, and 6. Thisapplies regardless of the breadth of the range.

REFERENCES

-   1. PTSD Statistics, PTSD United, available at    http://www.ptsuunited.org/ptsd-statistics-2/.-   2. American Psychiatric Association, Diagnostic and statistical    manual of mental disorders (5th ed.), Arlington, Va., American    Psychiatric Publishing (2013).-   3. AMPA receptor, Wikipedia,    https://en.wikipedia.org/wiki/AMA_receptor-   4. Park et al., Calcium-Permeable AMPA Receptors Mediate the    Induction of the Protein Kinase A-Dependent Component of Long-Term    Potentiation in the Hippocampus, Journal of Neuroscience 13 Jan.    2016, 36 (2) 622-631.-   5. Perusini, Jennifer Nicole, The Mechanisms of Fear Sensitization    Caused by Acute Traumatic Stress: from Induction to Expression to    Long-Lasting Reversal, UCLA: Psychology 0780 (2014), retrieved from:    http://escholarship.org/uc/item/3578829d.-   6. Perusini et al., Induction and Expression of Fear Sensitization    Caused by Acute Traumatic Stress, Neuropsychopharmacology, 41: 45-57    (2016).-   7. Rau, V., De Cola, J. P., & Fanselow, M. S. (2005). Stress-induced    enhancement of fear learning: An animal model of posttraumatic    stress disorder. Neuroscience & Biobehavioral Reviews, 29,    1207-1223.-   8. Rau, V. & Fanselow, M. S. (2009). Exposure to a stressor produces    a long lasting enhancement of fear learning in rats. Stress, 12,    25-33.-   9. Bolshakov et al., Different arrangement of hydrophobic and    nucleophilic components of channel binding sites in    N-methyl-D-aspartate and AMPA receptors of rat brain is revealed by    channel blockade, Neuroscience Letters 291 (2000) 101-104.-   10. Morris et al., Diagnostic accuracy of 18F amyloid PET tracers    for the diagnosis of Alzheimer's disease: a systematic review and    meta-analysis, Eur J Nucl Med Mol Imaging. 2016; 43: 374-385.-   11. Olah et al., Ionic Fluorination of Adamantane, Diamantane, and    Triphenylmethane with Nitrosyl Tetrafluoroborate/Pyridine    Polyhydrogen Fluoride (PPHF), J. Org. Chem., 1983, 48 (19), pp    3356-3358 (September 1983) (DOI: 10.1021/jo00167a050).-   12. Rozen and Gal, Direct Synthesis of Fluoro-Bicyclic Compounds    with Fluorine, J. Org. Chem., 1988, 53 (12), pp 2803-2807    (June 1988) (DOI: 10.1021/jo00247a026).

What is claimed is:
 1. A method of detecting Post-Traumatic Stress Disorder (PTSD) in a subject, comprising: a. administering to the subject a first amount of a radiolabeled composition comprising at least one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled with a radioactive isotope, wherein the radiolabeled composition comprises the following structure:

wherein the F is an [¹⁸F] radioisotope; b. creating at least one image of a brain of the subject using positron emission tomography (PET) or single-photon emission computed tomography (SPECT); and c. determining or quantifying from the at least one image a GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the brain of the subject based at least in part upon an amount of the radiolabeled composition detected in the at least one image, and comparing the density so determined or quantified with a predetermined baseline level, wherein a density greater than the predetermined baseline level indicates PTSD in the subject.
 2. A method of detecting GluA1 levels, or GluA1-containing, GluA2-lacking AMPAR levels, in an amygdala of a subject, comprising: administering to the subject a first amount of a radiolabeled composition comprising a ligand of GluA1 or a ligand of a GluA1-containing, GluA2-lacking AMPAR effective for detection of the radiolabeled composition in the amygdala of the subject using radiological imaging; and determining or quantifying, by the radiological imaging of the radiolabeled composition in the amygdala of the subject, GluA1 subunit density or GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the subject.
 3. A method of detecting Post-Traumatic Stress Disorder (PTSD) in a subject, comprising: administering to the subject a first amount of a radiolabeled composition comprising a ligand of GluA1 or a ligand of a GluA1-containing, GluA2-lacking AMPAR effective for detection of the radiolabeled composition in an amygdala of the subject using radiological imaging; and determining or quantifying, by the radiological imaging of the radiolabeled composition in the amygdala of the subject, GluA1 subunit density or GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the subject, and comparing the density so determined or quantified with a predetermined baseline level, wherein a density greater than the predetermined baseline level indicates PTSD in the subject.
 4. A method of detecting GluA1-mediated Post-Traumatic Stress Disorder (PTSD) in a subject, comprising: administering to the subject a first amount of a radiolabeled composition comprising a ligand of GluA1 or a ligand of a GluA1-containing, GluA2-lacking AMPAR effective for detection of the radiolabeled composition in an amygdala of a brain of the subject using radiological imaging; and determining or quantifying, by the radiological imaging of the radiolabeled composition in the amygdala of the subject, GluA1 subunit density or GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the subject, and comparing the density so determined or quantified with a predetermined baseline level, wherein a density greater than the predetermined baseline level indicates GluA1-mediated PTSD in the subject.
 5. The method of claim 2, 3, or 4, wherein determining or quantifying GluA1 subunit density or GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the subject comprises detecting or visualizing radioactive emissions of the radiolabeled composition in the amygdala after administration.
 6. The method of claim 2, 3, 4, or 5, wherein determining or quantifying GluA1 subunit density or GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the subject comprises determining or quantifying an amount of the radiolabeled composition in the amygdala after administration.
 7. The method of claim 2, 3, 4, 5, or 6, wherein the ligand comprises a ligand of GluA1.
 8. The method of claim 2, 3, 4, 5, or 6, wherein the ligand comprises a ligand of a GluA1-containing, GluA2-lacking AMPAR.
 9. The method of claim 2, 3, 4, 5, 6, 7, or 8, wherein the radiological imaging is positron emission tomography (PET) or single-photon emission computed tomography (SPECT) imaging.
 10. The method of claim 2, 3, 4, 5, 6, 7, 8, or 9, wherein the radiological imaging comprises using a portable electronic device to detect radiation levels associated with the radiolabeled composition.
 11. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, or 10, wherein the method further comprises comparing the radiolabeled composition in the amygdala after administration to a control amount.
 12. The method of claim 5, 6, 7, 8, 9, 10, or 11, wherein the method further comprises comparing receptor density associated with the radioactive emissions to a control receptor density.
 13. The method of claim 11 or 12, wherein the control amount or the control receptor density is a predetermined baseline level.
 14. A method of detecting GluA1 protein levels, or of GluA1-containing, GluA2-lacking AMPAR levels, in an amygdala of a subject comprising: a. administering to the subject a first amount of a radiolabeled composition comprising at least one ligand of GluA1 labeled with a radioactive isotope or comprising at least one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled with a radioactive isotope; b. creating at least one image of a brain of the subject using positron emission tomography (PET) or single-photon emission computed tomography (SPECT); and c. determining or quantifying from the at least one image a GluA1 subunit density or a GluA1-containing, GluA2-lacking AMPAR density in the amygdala of the brain of the subject based at least in part upon an amount of the radiolabeled composition detected in the at least one image, so as to detect GluA1 protein levels, or GluA1-containing, GluA2-lacking AMPAR levels, respectively, in an amygdala of a subject.
 15. A method of detecting Post-Traumatic Stress Disorder (PTSD) in a subject, comprising: a. administering to the subject a first amount of a radiolabeled composition comprising at least one ligand of GluA1 labeled with a radioactive isotope or comprising at least one ligand of a GluA1-containing, GluA2-lacking AMPAR labeled with a radioactive isotope; b. creating at least one image of a brain of the subject using positron emission tomography (PET) or single-photon emission computed tomography (SPECT); and c. determining or quantifying from the at least one image a GluA1 subunit density or a GluA1-containing, GluA2-lacking AMPAR density in an amygdala of the brain of the subject based at least in part upon an amount of the radiolabeled composition detected in the at least one image, and comparing the density so determined or quantified with a predetermined baseline level, wherein a density greater than the predetermined baseline level indicates PTSD in the subject.
 16. The method of claim 14 or 15 wherein the at least one image of the brain is created between 15 minutes and 3 hours following administration of the radiolabeled composition.
 17. The method of claim 14, 15, or 16, wherein the amount of the radiolabeled composition detected in the at least one image comprises an amount of radioactive emissions detected.
 18. The method of claim 14, 15, 16, or 17, further comprising determining whether the GluA1 subunit density or the GluA1-containing, GluA2-lacking AMPAR density in the amygdala exceeds a predetermined baseline level.
 19. The method of claim 18, further comprising providing a diagnosis for PTSD based at least in part upon the determination of whether the GluA1 subunit density or the GluA1-containing, GluA2-lacking AMPAR density in the amygdala exceeds the predetermined baseline level by at least a threshold amount.
 20. A method of treating Post-Traumatic Stress Disorder (PTSD) in a subject, comprising: a. receiving information indicating a detection of elevated levels of GluA1, either alone as a subunit protein or in GluA1-containing, GluA2-lacking calcium permeable AMPAR complexes in an amygdala of the subject, wherein the detection has been obtained by: i. administering to the subject an amount of a radiolabeled composition comprising at least one radiolabeled ligand of GluA1 or comprising at least one radiolabeled ligand of a GluA1-containing, GluA2-lacking AMPAR, the amount of the radiolabeled composition effective to detect the radiolabeled composition in the amygdala of the subject using positron emission tomography (PET) or single-photon emission computed tomography (SPECT); ii. determining or quantifying GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in the amygdala of the subject; and iii. determining that the GluA1 levels or the GluA1-containing, GluA2-lacking AMPAR levels exceed a predetermined baseline level; and b. administering to the subject an amount of a treatment composition effective to treat PTSD.
 21. The method of claim 20, wherein the respective amount of the treatment composition is effective to reduce GluA1 expression levels or GluA1-containing, GluA2-lacking AMPAR expression levels in the amygdala of a subject.
 22. The method of claim 21, wherein the treatment composition is selected from the group consisting of a nucleic acid, an antisense nucleic acid, a ribozyme, a peptide, a small molecule, an inhibitor of GluA1 expression or synthesis, an aptamer, and a peptidomimetic.
 23. The method of claim 20, wherein the respective amount of the treatment composition is effective to inhibit receptor function of GluA1-containing, GluA2-lacking AMPARs in the amygdala of the subject.
 24. The method of claim 23, wherein the treatment composition is selected from the group consisting of a peptide, a small molecule, an antagonist, an inhibitor, and a peptidomimetic.
 25. The method of claim 23 or 24, wherein the treatment composition comprises a GluA1-containing, GluA2-lacking AMPAR ligand.
 26. The method of claim 23, 24, or 25, wherein the treatment composition comprises an inhibitor of calcium permeable AMPAR function.
 27. The method of claim 13, 18, 19, 20, 21, 22, 23, 24, 25, or 26, wherein the predetermined baseline level is determined by: a. performing the imaging detection process on a plurality of subjects known not to be suffering from PTSD; b. determining respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the plurality of subjects; and c. computing as the predetermined baseline level a normalized average of the respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the plurality of subjects.
 28. The method of claim 13, 18, 19, 20, 21, 22, 23, 24, 25, or 26, wherein the predetermined baseline level is determined by: a. performing the imaging detection process on a first plurality of subjects known not to be suffering from PTSD; b. determining first respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the first plurality of subjects; c. computing a first normalized average of the first respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels; d. performing the imaging detection process on a second plurality of subjects known to be suffering from PTSD; e. determining second respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the second plurality of subjects; f. computing a second normalized average of the second respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels; and g. determining as the predetermined baseline level an amygdalar GluA1 level or GluA1-containing, GluA2-lacking AMPAR level based at least in part upon the first normalized average and the second normalized average.
 29. The method of claim 28, wherein the step of determining as the predetermined baseline level an amygdalar GluA1 level or GluA1-containing, GluA2-lacking AMPAR level based at least in part upon the first normalized average and the second normalized average comprises performing one or more statistical analyses.
 30. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, or 29, wherein the subject is a human subject.
 31. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30, wherein the radiolabeled composition comprises a detector of GluA1 expression, surface expression, AMPAR activity, or a combination thereof.
 32. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, or 31, wherein the radiolabeled composition is an inhibitor of AMPAR function.
 33. The method of claim 32, wherein the inhibitor is an inhibitor of calcium permeable AMPAR function.
 34. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, or 33, wherein the radiolabeled composition is selected from the group consisting of a nucleic acid, an antisense nucleic acid, a ribozyme, a peptide, a small molecule, an AMPAR antagonist, an aptamer, and a peptidomimetic.
 35. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, or 34, wherein the radiolabeled composition comprises a radioisotope.
 36. The method of claim 35, wherein the radioisotope is [¹⁸F].
 37. The method of claim 35, wherein the radioisotope is [¹¹C].
 38. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36, wherein the radiolabeled composition comprises N′-[(4-fluoro-1-adamantyl)methyl]pentane-1,5-diamine.
 39. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, or 38, wherein the radiolabeled composition comprises the following structure:

wherein the F is an [¹⁸F] radioisotope.
 40. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 37, wherein the radiolabeled composition comprises the following structure:

wherein the C is a [¹¹C] radioisotope.
 41. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 37, wherein the radiolabeled composition comprises the following structure:

wherein the C is a [¹¹C] radioisotope.
 42. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, or 41, wherein the radiolabeled composition comprises a saline or ethanol solution.
 43. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, or 42, wherein the radiolabeled composition is administered to the subject via injection into the bloodstream in a manner effective to enter the amygdala of the subject.
 44. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, or 42, wherein the radiolabeled composition is administered to the subject orally in a manner effective to enter the amygdala of the subject.
 45. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, or 42, wherein the radiolabeled composition is administered to the subject via inhalation.
 46. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, or 45, wherein the radiolabeled composition is administered to the subject via a nasal spray.
 47. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, or 46, wherein the first amount and/or the amount of the radiolabeled composition administered to the subject comprises a mass dose of the radiolabeled composition in a range of 0.0001 pg to 1 ng per kg of body weight of the subject.
 48. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, or 47, wherein an amount of radioactivity associated with the first amount and/or the amount of the radiolabeled composition administered to the subject falls in a range of 1 to 2000 MBq.
 49. The method of claim 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, or 48, wherein an effective radiation dose equivalent associated with the first amount and/or the amount of the radiolabeled composition administered to the subject falls in a range of 1 to 100 μSv.
 50. A composition comprising at least one radiolabeled detector of a GluA1-containing, GluA2-lacking calcium permeable (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor (AMPAR).
 51. The composition of claim 50, wherein the GluA1-containing, GluA2-lacking calcium permeable AMPAR is a human AMPAR.
 52. The composition of claim 50 or 51, wherein the radiolabeled detector comprises a ligand of a GluA1-containing, GluA2-lacking calcium permeable AMPARs.
 53. A composition comprising at least one radiolabeled detector of GluA1 protein.
 54. The composition of claim 50, 51, 52, or 53, wherein the radiolabeled detector is a detector of GluA1 expression, surface expression, AMPAR activity, or a combination thereof.
 55. The composition of claim 50, 51, 52, 53, or 54, wherein the radiolabeled detector is selected from the group consisting of a nucleic acid, an antisense nucleic acid, a ribozyme, a peptide, a small molecule, an AMPAR antagonist, an aptamer, and a peptidomimetic.
 56. The composition of claim 50, 51, 52, 53, 54, or 55, wherein the radiolabeled detector comprises a radioisotope.
 57. The composition of claim 56, wherein the radioisotope is [¹⁸F].
 58. The composition of claim 50, 51, 52, 53, 54, 55, 56, or 57, wherein the radiolabeled detector comprises N′-[(4-fluoro-1-adamantyl)methyl]pentane-1,5-diamine.
 59. The composition of claim 50, 51, 52, 53, 54, 55, 56, 57, or 58, wherein the radiolabeled detector comprises the following structure:

wherein the F is an [¹⁸F] isotope.
 60. The composition of claim 56, wherein the radioisotope is [¹¹C].
 61. The composition of claim 50, 51, 52, 53, 54, 55, 56, or 60, wherein the radiolabeled detector comprises the following structure:

wherein the C is a [¹¹C] isotope.
 62. The composition of claim 50, 51, 52, 53, 54, 55, 56, or 60, wherein the radiolabeled detector comprises the following structure:

wherein the C is a [¹¹C] isotope.
 63. The composition of claim 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, or 62, wherein the composition comprises a saline solution of the radiolabeled detector.
 64. The composition of claim 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, or 62, wherein the composition comprises an ethanol solution of the radiolabeled detector.
 65. The composition of claim 64, wherein the ethanol is diluted with water or saline.
 66. The composition of claim 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, or 65, wherein the composition is suitable as a radiolabeled tracer in an imaging detection process.
 67. The composition of claim 66, wherein the imaging detection process is positron emission tomography (PET) or single-photon emission computed tomography (SPECT).
 68. The composition of claim 66 or 67, wherein the imaging detection process is used to quantify or determine expression of GluA1 protein or of GluA1-containing, GluA2-lacking AMPARs in an amygdala of a subject.
 69. The composition of claim 68, wherein the imaging detection process is used to determine whether the GluA1 protein or the GluA1-containing, GluA2-lacking AMPARs exceed a predetermined baseline level.
 70. The composition of claim 68 or 69, wherein the imaging detection process is used to determine an amount by which the GluA1 protein or the GluA1-containing, GluA2-lacking AMPARs exceed a predetermined baseline level.
 71. The composition of claim 69 or 70, wherein the predetermined baseline level is determined by: a. performing the imaging detection process on a plurality of subjects known not to be suffering from PTSD; b. determining respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels, in each of the plurality of subjects; and c. computing as the predetermined baseline level a normalized average of the respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the plurality of subjects.
 72. The composition of claim 69 or 70, wherein the predetermined baseline level is determined by: a. performing the imaging detection process on a first plurality of subjects known not to be suffering from PTSD; b. determining first respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the first plurality of subjects; c. computing a first normalized average of the first respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels; d. performing the imaging detection process on a second plurality of subjects known to be suffering from PTSD; e. determining second respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels in each of the second plurality of subjects; f. computing a second normalized average of the second respective amygdalar GluA1 levels or GluA1-containing, GluA2-lacking AMPAR levels; and g. determining as the predetermined baseline level an amygdalar GluA1 level or GluA1-containing, GluA2-lacking AMPAR level based at least in part upon the first normalized average and the second normalized average.
 73. The composition of claim 72, wherein the step of determining as the predetermined baseline an amygdalar GluA1 level or GluA1-containing, GluA2-lacking AMPAR level based at least in part upon the first normalized average and the second normalized average comprises performing one or more statistical analyses.
 74. The composition of claim 66, 67, 68, 69, 70, 71, 72, or 73, wherein the imaging detection process is usable to detect post-traumatic stress disorder in a subject.
 75. A compound having the following structure:

wherein the F is an [¹⁸F] radioisotope.
 76. A compound having the following structure:

wherein the C is a [¹¹C] radioisotope.
 77. A compound having the following structure:

wherein the C is a [¹¹C] radioisotope.
 78. A method of synthesizing a radiolabeled compound comprising performing a radiofluorination reaction on a first compound having the following structure:

so as to produce a second compound having the following structure:

wherein the F is an [¹⁸F] radioisotope.
 79. A method of synthesizing a radiolabeled compound comprising: a. obtaining an amount of [¹⁸F]; b. eluting the [¹⁸F] with a phase transfer catalyst KF/K2.2.2 so as to produce a solution of [¹⁸F]KF/K2.2.2 complex; c. adding a first compound having the following structure:

to the solution of [¹⁸F]KF/K2.2.2 complex so as to perform a radiofluorination reaction to produce a second compound having the following structure:

wherein the F is an [¹⁸F] radioisotope.
 80. The method of claim 79, further comprising purifying the reaction contents from step (c) to produce a purified compound.
 81. The method of claim 80, wherein purifying the reaction contents comprises performing radio-HPLC on the product of the radiofluorination reaction.
 82. A method of producing a radiolabeled compound comprising: a. synthesizing a [¹⁸F]fluoride solution comprising [¹⁸F]fluoride via a (p,n) reaction of [¹⁸O]water by proton bombardment in a cyclotron; b. extracting the [¹⁸F]fluoride by passing the [¹⁸F]fluoride solution through a preconditioned QMA anion exchange cartridge; c. eluting, in a reaction vessel, the extracted [¹⁸F]fluoride with a first solvent comprising an acetonitrile (MeCN)/water solution containing a phase transfer catalyst KF/K2.2.2; d. heating the reaction vessel; e. applying a vacuum to the reaction vessel so as to remove the first solvent and leave a dried residue of a [¹⁸F]KF/K2.2.2 complex; f. adding, to the reaction vessel, MeCN to re-dissolve the dried residue; g. evaporating, via application of heat and vacuum to the reaction vessel, so as to remove residual water via azeotropic distillation leaving a second dried residue of [¹⁸F]KF/K2.2.2 complex; h. mixing, in a second solvent in the reaction vessel, the second dried residue of [¹⁸F]KF/K2.2.2 complex and a compound having the following structure:

i. heating the reaction vessel so as to perform a radiofluorination reaction to produce a radiolabeled compound having the following structure:

wherein the F is an [¹⁸F] radioisotope; and j. purifying, via radio-High Performance Liquid Chromatography (HPLC), the radiofluorination reaction contents from step (i) to produce a purified radiolabeled compound.
 83. The method of claim 82, further comprising formulating the purified radiolabeled compound in saline.
 84. The method of claim 82 or 83, wherein the second solvent comprises MeCN or dimethyl sulfoxide (DMSO). 